The cerebral cavernous malformation (CCM) is a common vascular anomaly, predisposing to a lifetime risk of stroke and other neurologic sequelae. Lesions occur in either a sporadic form or in an autosomal- dominant inherited form, the latter due to mutation in one of three genes. Molecular genetic analyses of surgically resected CCM lesions by the Awad and Marchuk laboratories has uncovered second-hit somatic CCM gene mutations in endothelial cells lining the vascular caverns, suggesting a two-hit mutational mechanism of CCM pathogenesis. Using this knowledge, we have developed robust animal models of CCM recapitulating the histology, molecular signatures and ultrastructure of the human lesions. Although we can now describe the major stages of lesion pathogenesis, the underlying molecular switches that modulate the progression of these stages remain unknown. In parallel work, the Ginsberg and other laboratories have shown that loss of CCM gene function impairs endothelial cell junctions, in part regulated by RhoA/ROCK activity. Yet, the Ginsberg, Kahn, and other laboratories have shown that loss of CCM function alters other major signaling pathways such as Notch, Wnt/-catenin, FOXO1, and KFL2/MEKK2 signaling. The centrality of RhoA/ROCK activity in CCM pathogenesis, and hence its optimal therapeutic target(s), remain unknown. Our central hypothesis of this P01 proposal is that the loss of CCM proteins contributes to lesion formation via multiple aberrant signaling pathways, some of which are RhoA/ROCK-independent. We further propose that different signaling and genetic aberrations modulate distinct stages of lesion development and maturation. We propose to analyze molecular genetic events during lesion development, and investigate associated signaling in vivo and in vitro. Our murine models enable us to investigate the role of these pathways in vivo at the different stages of CCM pathogenesis, and our collection of surgically resected CCMs allows us to validate these findings in the clinically relevant mature human lesion. The continuum of in vitro, in vivo and detailed analysis of mouse and human lesions will help us create an ordered scheme of aberrant signaling networks in relation to lesion pathogenesis, and translate new fundamental insights into rational therapeutic strategies for this disease.